Our long-term objective is to characterize the ways in which the responsiveness of airway smooth muscle to relaxant and contractile stimuli may be regulated in health and disease. Because of recent demonstrations that smooth muscle from the airways of asthmatic patients has diminished beta-adrenergic relaxant responsiveness (Am Rev Respir Dis 134:57-61, 1986), the specific aim of this proposal is to characterize selected experimental interventions which reduce beta-adrenergic responses in the trachealis muscle of dogs in vivo and in vitro and in cultured tracheal smooth muscle cells. We will focus our studies on selected components of the beta-adrenergic cascade where changes in the muscle cell may alter responses potently. These components will be beta-adrenergic receptors and the guanyl nucleotide regulatory proteins (Ns and Ni) which control the activity of adenylate cyclase. New techniques will be used to localize the receptors ultrastructurally in the intact muscle and to characterize Ns and Ni in subcellular fractions isolated from the muscle. Three of our specific hypotheses will be that: (1) brief exposure of the muscle to isoproterenol reduces its beta- adrenergic responsiveness because of specific changes in the receptor, which include its early uncoupling from Ns, its subsequent internalization into smooth muscle cells, and its eventual degradation in smooth muscle cells; (2) prolonged exposure of the muscle to prostaglandin E-2 reduces its beta-adrenergic responsiveness because prostaglandin E-2 activates cyclic AMP-dependent protein kinase, which in turn alters Ns or Ni; (3) exposure of the muscle to products released from purified canine mastocytoma cells reduces the muscle's beta-adrenergic responsiveness, because proteases released from the mastocytoma cells alter receptors, Ns, or Ni in the muscle and because other masto-cytoma cell mediators (such as histamine) activate protein kinase C, which in turn alters receptors, Ns, or Ni in the muscle. We believe that these findings will provide important new information regarding the mechanisms by which beta-adrenergic relaxant responses are reduced in the airways of asthmatic patients.
